人牙周膜干细胞的分离鉴定及骨形态发生蛋白-2对其趋化效应的研究
Culturing and characterization of human periodontal ligament stem cells and investigating their chemotactic responses to bone morphogenetic protein-2
作者：杜令倩 杨丕山 赵宁 葛少华
Author：Du Lingqian1，2, Yang Pishan1，3, Zhao Ning1，3, Ge Shaohua1，3
收稿日期：2012-02-25 年卷（期）页码：2012,30(1):13-13-17
期刊名称：华西口腔医学杂志
Journal Name：West China Journal of Stomatology
关键字：骨形态发生蛋白-2,牙周膜干细胞,趋化作用,
Key words：bone morphogenetic protein-2,periodontal ligament stem cells,chemotaxis,
基金项目：
国家自然科学基金资助项目（81100756）；山东省自然科学基金资助项目（ZR2009CM118）；山东大学自主创新基金资助项目（2009TS047）；山东省科技攻关计划基金资助项目（2010GSF10248）
中文摘要
目的研究人牙周膜干细胞（PDLSCs）对骨形态发生蛋白-2（BMP-2）的趋化反应。方法通过有限稀释法分离、培养人PDLSCs，利用免疫荧光染色检测人PDLSCs波形丝蛋白及干细胞表面标志物STRO-1的表达，检测人PDLSCs多向分化能力，通过克隆形成实验和5-溴-2-脱氧尿嘧啶核苷（BrdU）共培养的方法检测其干细胞特性。利用24孔的Transwell细胞培养室来检测人PDLSCs对BMP-2的趋化反应，光镜下计迁移至滤膜下侧面的不同视野的细胞数。结果人PDLSCs抗波形丝蛋白染色阳性，表达干细胞表面标志物STRO-1，体外诱导培养的人PDLSCs能够向成骨细胞和成脂细胞分化，具有较高的自我更新能力，并在体外呈克隆状生长。在100、200 ng·mL-1 BMP-2实验组，Transwell细胞培养室中迁移的细胞数目显著多于空白对照组（P
英文摘要
Objective To investigate the chemotactic response of human periodontal ligament stem cells（PDLSCs）to bone morphogenetic protein-2（BMP-2）. Methods Human PDLSCs were obtained from clinically healthy premolars extracted for orthodontic reasons and used to isolate PDLSCs by limited dilution method. The expression of Vimentin and stem cell marker STRO-1 on PDLSCs were demonstrated with immunocytochemical staining. Differentiation assay was used to detect the differentiation potential of PDLSCs. Cloning formation experiment and 5-bromo-2-deoxyuridine （BrdU） incorporation assay were used to determine the stem cell characteristics of PDLSCs. The chemotactic effect of BMP-2 on PDLSCs was detected by using a 24-multiwell Transwell cell culture chamber. The number of net migrated cells was counted in different microscope fields. Results Human PDLSCs displayed positive staining for Vimentin and expressed the stem cell marker STRO-1. These cells differentiated into osteoblasts and adipocytes under defined culture conditions, possessed high self-renewal potential and formed single-cell colonies in vitro. The number of cells migrating at concentrations of 100, 200 ng·mL-1 of BMP-2 in Transwell cell culture chamber was significantly higher than that of negative control （P
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